Abstract Telomeres function as an aging clock because most human somatic cells lack telomerase expression and their telomeres progressively shorten upon successive cell divisions. Accordingly, telomere shortening is a critical factor of human aging and telomerase activation is essential for the development of many human cancers. On the other hand, cells from some other organisms, including mice, do not exhibit telomere-mediated replicative aging. Mice possess long heterogeneous telomeres and ubiquitous telomerase activities in somatic tissues. This inter-species difference has become a bottleneck for addressing many fundamental questions in human aging and cancer biology using mouse models. Thus, the long-term goal of this project is to genetically engineer mouse strains with human-like telomere homeostasis and to test the hypothesis that short telomeres and repressed telomerase confer replicative aging in mice. Our recent studies have identified several distal regulatory elements critical for the repression of hTERT gene. In this proposal, we will test the hypothesis that incorporation of these elements into their corresponding mouse genomic sites leads to human-like TERT gene regulation in mice. We will first engineer mouse embryonic stem cells (ESCs) with a humanized mTERT locus (hmTERT) and then use these ESCs to generate mouse strains with human-like telomerase regulation. The regulation of hmTERT locus will be determined both during in vitro ESC differentiation and during mouse development. We anticipate that these mice will serve as an improved model for studying human aging and cancer.